Subject(s)
Coordination Complexes , Copper/chemistry , DNA Topoisomerases, Type I/chemistry , Topoisomerase I Inhibitors , Zinc/chemistry , Catalytic Domain , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , Humans , Topoisomerase I Inhibitors/chemical synthesis , Topoisomerase I Inhibitors/chemistryABSTRACT
The ability of oxindolimine copper(II) and zinc(II) complexes, known to have antitumor activity, to inhibit human topoisomerase IB has been tested through enzymatic kinetic assays and molecular docking simulations. These copper and zinc compounds are able to inhibit remarkably the cleavage reaction and only partially the religation step, the copper compound being more efficient than the zinc one. A complete inhibition activity of the cleavage is only obtained when the enzyme is pre-incubated with the compound, the inhibition being irreversible and reversible for the copper and zinc compounds, respectively. The relative stability of such complexes was estimated by competitive equilibria with human serum albumin (HSA), monitored by CD spectroscopy. The copper species shows a log KCuL = 17.2, while the analogous zinc complex exhibits a log KZnL = 7.2. Molecular docking simulation studies show that the almost square planar geometry of the copper compound allows a direct coordination of the metal with two amino acids (Glu492, Asp563) of the enzyme at variance of the zinc compound which has a more tetrahedral geometry. Altogether, the data indicate that the different coordination geometry achieved by the two transition metal ions has an important role in modulating their efficiency as topoisomerase I inhibitors.
Subject(s)
Bridged Bicyclo Compounds/chemistry , Coordination Complexes/chemistry , Copper/chemistry , DNA Topoisomerases, Type I/metabolism , Zinc/chemistry , Aspartic Acid/chemistry , Aspartic Acid/metabolism , Binding Sites , Biocatalysis/drug effects , Bridged Bicyclo Compounds/metabolism , Circular Dichroism , Coordination Complexes/metabolism , Coordination Complexes/pharmacology , Copper/metabolism , DNA Topoisomerases, Type I/chemistry , DNA, Circular/chemistry , DNA, Circular/genetics , DNA, Circular/metabolism , Glutamic Acid/chemistry , Glutamic Acid/metabolism , Humans , Kinetics , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Structure , Oligonucleotides/chemistry , Oligonucleotides/genetics , Oligonucleotides/metabolism , Protein Binding , Protein Structure, Tertiary , Topoisomerase I Inhibitors/chemistry , Topoisomerase I Inhibitors/metabolism , Topoisomerase I Inhibitors/pharmacology , Zinc/metabolismABSTRACT
Topoisomerases IB are anticancer and antimicrobial targets whose inhibition by several natural and synthetic compounds has been documented over the last three decades. Here we show that kakuol, a natural compound isolated from the rhizomes of Asarum sieboldii, and a derivative analogue are able to inhibit the DNA relaxation mediated by the human enzyme. The analogue is the most efficient one and the inhibitory effect is enhanced upon pre-incubation with the enzyme. Analysis of the different steps of the catalytic cycle indicates that the inhibition occurs at the cleavage level and does not prevent DNA binding. Molecular docking shows that the compound preferentially binds near the active site at the bottom of the catalytic residue Tyr723, providing an atomistic explanation for its inhibitory activity.
Subject(s)
Benzodioxoles/chemistry , Benzodioxoles/pharmacology , Biological Products/chemistry , Biological Products/pharmacology , DNA Topoisomerases, Type I/metabolism , DNA/chemistry , Nucleic Acid Conformation/drug effects , Propiophenones/chemistry , Propiophenones/pharmacology , Base Sequence , Catalytic Domain , DNA/genetics , DNA/metabolism , DNA Topoisomerases, Type I/chemistry , Humans , Models, Molecular , Topoisomerase I Inhibitors/chemistry , Topoisomerase I Inhibitors/pharmacologyABSTRACT
The different steps of the topoisomerase I catalytic cycle have been analyzed in the presence of the plant alkaloid thaspine (1- (2-(Dimethylamino)ethyl)-3,8-dimethoxychromeno[5,4,3-cde]chromene-5,10-dione), known to induce apoptosis in colon carcinoma cells. The experiments indicate that thaspine inhibits both the cleavage and the religation steps of the enzyme reaction. The inhibition is reversible and the effect is enhanced upon pre-incubation. Molecular docking simulations of thaspine over topoisomerase I, in the presence or absence of the DNA substrate, show that thaspine, when interacting with the enzyme alone in the closed or in the open state, can bind in proximity of the active residues preventing the cleavage reaction, whilst when docked with the enzyme-DNA cleavable complex intercalates between the DNA bases in a way similar to that found for camptothecin, explaining its religation inhibition. These results unequivocally demonstrate that thaspine targets human topoisomerase I .
Subject(s)
Alkaloids/pharmacology , Antineoplastic Agents/pharmacology , Biological Products/pharmacology , DNA Topoisomerases, Type I/metabolism , Topoisomerase I Inhibitors/pharmacology , Alkaloids/chemistry , Antineoplastic Agents/chemistry , Biological Products/chemistry , DNA Topoisomerases, Type I/genetics , DNA Topoisomerases, Type I/isolation & purification , Humans , Kinetics , Models, Molecular , Molecular Structure , Structure-Activity Relationship , Topoisomerase I Inhibitors/chemistryABSTRACT
A gold(III) compound [Au(C^N^C)(IMe)]CF(3)SO(3) (Gold III) has been reported to have anticancer properties as it is able to reduce topoisomerase IB activity in vitro and suppress tumor growth in nude mice model. Here we have investigated the mechanism of inhibition of human topoisomerase IB activity by this compound, analyzing the various steps of the catalytic cycle. DNA supercoiled relaxation and the cleavage reaction are inhibited, but Gold III does not perturb the religation reaction, in contrast to what has been observed for camptothecin. Pre-incubation of enzyme with the inhibitor before adding DNA substrate increases the inhibitory effect. In addition, when Gold III is preincubated with the enzyme it prevents the stabilization of the cleavable complex by camptothecin. The analysis of the DNA-topoisomerase binding reaction indicates that the compound acts as a topoisomerase I inhibitor by preventing the enzyme-DNA interaction.
